Adenosine Deaminase

Structure, Function (General Info)

Article talked about the adenosine deaminase binding protein, known as type II transmembrane serine protease dipeptidyl peptidase IV, and studies on DPPIV binding. It was revealed that the N-linked glycosylation on Asn residues in DPPIV interacts with adenosine deaminase. The N-linked glycosylation contribute to binding with the ADA protein but does not contribute to peptidase activity of DPPIV. This article is helpful in terms of understanding how adenosine deaminase binds with other proteins for further stimulation.

Aertgerrts et al. investigated the structure of human adenosine deaminase ADA2 through protein models and enzymatic kinetics with wild and mutated protein. Crystalized structures were used for model accuracy. ADA2 revealed to be more complex than ADA1 with only a monomeric single domain. Figures in paper showed structures of human adenosine deaminase and could be used for reference in determining how ADA minds with gold nanoparticles.

Sauer et al. provided an overview of Adenosine deaminase cause for severe combined immunodeficiency (ADA-SCID) and patterns before and after treatments in patients. Immune dysregulation are related to alterations in purine metabolism that is caused by the lack of ADA. Article help explain the importance of adenosine deaminase in cellular signaling and highlighted descreased immune functions when adenosine deaminase is lacking.

Activity Assays

The conversion of adenosine to inosine by adenosine deaminase was spectroscopically analyzed at 265 nm for a period of 15 minutes. The number of assays that could be conducted at one time was increased by carrying out analysis in 96-well microplates. This assay also varies from the standard procedure in that the reaction time was decreased from 60 minutes.

Aptamer-based sensor is reported in this article for time-resolved fluorescence assay of adenosine deaminase. The aptasensor works by binding to DNA strands which interacts with adenosine deaminase, receiving a strong signal. This could be potential jumpoff point to design experiments for fluorscence assay for adenosine deaminase activity.

Bovine Serum Albumin

Structure, Function (General Info)

Ahmed et al. located binding sites of lead, Pb(II) with human serum and bovine serum albumins(BSA) at physiological conditions. Through the use of UV-visible, fluorescence, and X-ray photoelectron spectroscopics, Pb binding sites were found and analyzed. Pb bound is 0.7 per HSA and BSA complex. Article provide protocol in measure concentration of bound BSA under UV-visible and fluorescence spectroscopic, which could be used for reference.

Interaction with other nobel metals

Silva et al. studied the binding of chlorpromazine and hemin to bSA via the use of fluorescence quenching technique. A range from 300 to 450nm has shown to be the emission spectra for each quencher addition to BSA. Primary binding site for hemin to BSA was found in this article between the two tryptophan residue in BSA protein. This could help in understanding how BSA binds to other molecules and help in understanding the structure of BSA itself.

Tests the biofunctionalization of silver nano-particles and as a result investigates the interaction between AgNPs and BSA at different concentrations as observed by a UV-Vis.

Discovered that AgNPs coated in BSA were observed under UV-Vis to be quite different than the uncoated particles. The BSA adsorption on the surface prevented the Ag nanoparticles from aggregating in solutions of pH greater than 5 which may prevent the uptake of the NPs in live cells.

Lysozyme

Structure, Function (General Info)

information about lysozyme -- "Lysozyme is a globular protein with a molecular weight of approximately 14.7 kDa. Lysozyme has been used as a model protein for this study, since its biophysical properties are well understood and the specific contribution of the sugar to its stability over a range of solution conditions can easily be compared with that of the native protein. Lysozyme is a widely studied protein due to its rich phase behavior(17, 18) including crystallization,(19) liquid–liquid phase separation,(20-22) and the formation of equilibrium clusters(23-25) and gels.(24) The solution behavior of lysozyme is also well understood in terms of its interaction with salt ions.(26-28)" from James and McManus J. Phys. Chem. B, 2012, 116 (34), pp 10182–10188

Article looked at domains of lysozyme and determined the optimal enzymatic conditions for activity for highly thermostable lysoenzyme. The thermostability can be applied for maximum reactivity when conducting assays on lysozyme. Furthermore, Enzymatic activity were quantified through the use of mass spectroscopy. Protocal could be considered for replication.

A novel technique of unfolding and then refolding lysozyme was investigated and analyzed using spectroscopy, circular dichroism, and activity assay for lysozyme. The lysozyme was first precipitated out of solution by adding a negatively-charged surfactants, then dissolved by adding a positively-charged surfactant. Upon its dissolution, lysozyme was observed to spontaneously refold to its native state.

The anionic/cationic surfactant pairs studied all produced similar results in that the amount of lysozyme that refolded was observed to be the same in each surfactant pair.

Horseradish Peroxidase

Structure, Function (General Info)

Lysine modification in horseradish peroxidase decreases stability of the protein when the protein is unfolded. Under 60°C, the structure of horseradish peroxidase becomes less compact. This brings insights to performing the reaction between gold nanoparticles with the enzyme under the ideal temperatures.

Horseperoxidase catalyze the reduction of H2O2. Biosensor was developed to measure the concentration of horseradish peroxidase in aqueous solution as gelation reagent to immobolize nanocomposite onto electrode surface. Furthermore, UV-vis and FT-IR spectroscopic analysis of HRP among other enzymes were made. This data could be used for reference during HRP assay tests.

The horseradish peroxidase assay using luminol and enhanced by p-iodophenol was optimized for determining the presence and concentration of H2O2 in solution. The intensity of the chemiluminescence was found to be optimal at pH 8.5. The study outlines a protocol that is less vague than others and also includes calibration curves.

Protein/Peptide Nanoparticle Synthesis

Bovine Serum Albumin

Mathematical models were used to determine the effects of reaction conditions, including pH, concentration, temperature, and solvent addition rate, on the formation of BSA nanoparticles. The BSA NPs were synthesized by coacervation, then crosslinked with glutaraldehyde. The study indicated that glutaraldehyde did not have an effect on the size of the NPs, stabilized the BSA NPs, and decreased swelling in water.

Silk fibroin nanoparticles were synthesized by first dissolving silk fibers in CaCl2, then adding water-miscible organic solvents. Analysis of the NPs indicated that they were insoluble in water and that their morphology depended on the types of organic solvents used. The study indicated that acetone produced NPs with the most globular shape.

AuNPs are synthesized using sonic vibration, with sodium citrate dihydrate as a stabilizing and reducing agent. Compared to previous protocols, this method takes an extremely short amount of time to prepare AuNPs and does not require the solutions to be heated.

Extract from Zingiber officinale, a type of ginger, is used as a stabilizing and reducing agent to synthesize AuNPs. According to the literature, ginger extract is more efficient than aspirin in decreasing blood platelet aggregation, so this method of AuNP synthesis is promising for use in medical treatments.

Explored the possibility of coating Gold nano-particles in cinnamon to aid in cancer detection procedures. The alcoholic components of cinnamon reduce NaAuCl4 to produce gold- nanoparticles. the Cin-AuNPs are biocompatible and can serve as excellent CT/ photoacoustic contrast-enhancement agents to detect tumors.

DNA-modified gold nanoparticles were used to detect single-base subsitutions in DNA by colorimetry/spectroscopy. The nanoparticles were stabilized with single-stranded DNA molecules. AuNPs aggregate by crosslinking with DNA molecules but also without crosslinking (NCL). The NCL aggregations were able to detect single-base mutations at very low concentrations.

A Dendronization strategy was used to control the interparticle spacing and optical properties of gold nanoparticle aggregates in aqueous solution. Tests how disulﬁde concentration, temperature, time and nature of the ligand (dendritic vs nondendritic), determine the control exerted over the size and stability of the NP aggregates.

Observed the photoluminescence of water-soluble gold nano-particles which were wrapped in BSA protective layer. Alternative to organic fluorophores or semiconductor nanoparticles for biological labeling and imaging.

The size effects and contributions from surface characteristics of small nano particles.

helps determine which combination is most negatively charged or positively charged.

Isothermal titration calorimetry (ITC) measures the heats of interaction of reactants and therefore can determine strengths of bonds between the reactants and help deduce how the reactants are bonded together.

Aggregation Stability

The study discovered that gold nanoparticles cause BSA proteins to become partially unfolded at the nanoparticle-protein interface. Aggregation of proteins is catalyzed by these misfolded BSA proteins.